Genetically induced alterations in the cellular apoptotic response are important for mechanisms of environmental carcinogenesis. Tumor suppressor genes may function to regulate proliferative or apoptotic signaling pathways. The functions of the Tsc2 tumor suppressor gene are unknown. The Tsc2 gene product, tuberin, has been shown to have GTPase activating protein (GAP) activity that may regulate apoptotic signaling pathways that depend on GTPase activity. The Raf-1/ERK and PI3 kinase/Akt signaling pathways are two signaling pathways that may depend on GTPase activity. The proposed studies are designed to determine if Tsc2 functions to regulate apoptotic signaling and whether it does so by altering the activity or expression of signal transduction proteins in these pathways, specifically Raf- 1, ERK, and Akt. To test this hypothesis, we will specifically: (1) measure the effect of Tsc2 expression on the apoptotic response of renal epithelial cells exposed to apoptosis-inducing compounds; (2) develop stable lines of Tsc2 null cells that have inducible and readily detectable expression of wild type Tsc2 or the Tsc2 GAP domain and measure the apoptotic response of these cells to apoptosis inducing compounds; and (3) examine the effect of wild type Tsc2 and Tsc2 GAP domain expression on Raf-1, ERK and Akt levels and activities. The long-term objective of these studies is to establish a mechanistic link between gene expression, apoptosis, environmental exposure, and carcinogenesis. The proposed studies may be relevant to future studies that seek to predict the influence of environmental factors on the onset of diseases with genetic predisposition. These endpoints may also be useful in the development of new anti-neoplastic strategies.